Sexual transmission remains the dominant mode of HIV transmission. Young women in sub-Saharan Africa bear a disproportionate burden of HIV infection, underscoring an urgent need for women-initiated prevention technologies. The CAPRISA 004 trial demonstrated that 1% tenofovir gel used before and after sex can reduce HIV infection rates by 39%, and 54% in women who used tenofovir gel consistently. This trial represents the highest efficacy for topically-applied ARVs achieved to date, and provides a strong foundation for increasing efforts to enhance the efficacy of ARV-based microbicides. Data from CAPRISA 004 demonstrates compellingly that mucosal inflammation is an independent predictor of HIV acquisition in this trial. Further, participants with tenofovir drug levels >1000ng/ml had significantly higher levels of protection than those with lower levels, suggesting some intracellular competition for the phosphates necessary for tenofovir's antiviral activity. Here we propose to expand this observation using a combination of clinical trial samples, and in vitro and in vivo models to test the hypothesis that a certain threshold of mucosal inflammation in the genital tract is required for HIV to establish infection, and that women who become infected despite using the protective tenofovir gel will display the most extreme genital tract inflammatory phenotype. Understanding the nature, mechanism and magnitude of this effect could have important implications for HIV prevention strategies that block genital inflammation. We have assembled an excellent research team with substantial expertise in mucosal immunology, clinical trials and epidemiology of HIV, and appropriate experimental systems to study HIV replication and immune activation in vitro, and in vivo in the emerging humanized mouse model. We have also established laboratory methods and generated preliminary data to study the link between inflammation and the presence of HIV target cells in the female genital tract. This is central to addressing our proposed mechanism of tenofovir metabolite-dATP substrate competition in the context of inflammation and activated HIV target cell recruitment. This proposal builds on a model of heterogeneity in susceptibility to HIV infection, and adds an important dimension by conducting a comprehensive cohort study of the completed CAPRISA 004 and ongoing CAPRISA 008 implementation trials of 1% tenofovir gel. Enhanced HIV prevention modalities for women remains an urgent public health imperative in Southern Africa and globally.